LOCUS       sequence-436020-        5211 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for ceramide (putative, mammalian expression).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5211)
  AUTHORS   Girik V, van Ek L, Dentand Quadri I, Azam M, Cruz Cobo M, Mandavit 
            M, Riezman I, Riezman H, Gavin AC, Nunes-Hasler P
  TITLE     Development of Genetically Encoded Fluorescent KSR1-Based Probes to 
            Track Ceramides during Phagocytosis.
  JOURNAL   Int J Mol Sci. 2024 Mar 5;25(5):2996. doi: 10.3390/ijms25052996.
  PUBMED    38474242
REFERENCE   2  (bases 1 to 5211)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Int J Mol 
            Sci."; date: "2024-03-5"; volume: "25(5):2996. doi"; pages: " 
            10.3390/ijms25052996"
COMMENT     Sequence Label: pEGFP-SET-EMD (N-SET-EMD)
FEATURES             Location/Qualifiers
     source          1..5211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1999..2120
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2036..2055)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2090..2109
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2127..2582)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2264..2285)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2476..2495
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2609..2713
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2715..3072
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2715..2906
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2738..2758)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2923..3058
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3107..3901
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3107..3901
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3161..3180)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3771..3790
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4089..4108)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4133..4180
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4509..5097
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4998..5017
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct ggcggttcct ctggtggtgg tggtgcgcag
     1381 aagaggtcag aattgatcgc caaaatccca aatttttggg taacaacatt tgtcaaccat
     1441 ccacaagtgt ctgcactgct tggggaggaa gatgaagagg cactgcatta tttgaccaga
     1501 gttgaagtga cagaatttga agatattaaa tcaggttaca gaatagattt ttattttgat
     1561 gaaaatcctt actttgaaaa taaagttctc tccaaagaat ttcatctgaa tgagagtggt
     1621 gatccatctt cgaagtccac cgaaatcaaa tggaaatctg gaaaggattt gacgaaacgt
     1681 tcgagtcaaa cgcagaataa agccagcagg aagaggcagc atgaggaacc agagagcttc
     1741 tttacctggt ttactgacca ttctgatgca ggtgctgatg agttaggaga ggtcatcaaa
     1801 gatgatattt ggccaaaccc attacagtac tacttggttc ccgatatgga ttaaggtacc
     1861 gcgggcccgg gatccaccgg atctagataa ctgatcataa tcagccatac cacatttgta
     1921 gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg
     1981 aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     2041 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     2101 aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc
     2161 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     2221 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     2281 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     2341 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     2401 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     2461 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     2521 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     2581 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     2641 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     2701 aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt
     2761 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     2821 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     2881 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     2941 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3001 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3061 ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     3121 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     3181 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     3241 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc
     3301 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     3361 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     3421 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     3481 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     3541 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     3601 cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt
     3661 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     3721 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     3781 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     3841 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     3901 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     3961 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4021 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg
     4081 aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata
     4141 aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc
     4201 cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt
     4261 tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac
     4321 gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta gattgattta
     4381 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     4441 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     4501 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     4561 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     4621 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     4681 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     4741 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     4801 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     4861 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     4921 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     4981 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     5041 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     5101 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     5161 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgccatgca t
//
